Recent evidence has been provided that thyroid hormones may also play a role in carcinogenesis, and pioneering investigations in this area have demonstrated that the thyroid hormones such as 3,5,3'-triiodo-L-thyronine (T3) and L-Thyroxine (T4) can stimulate cancer cell proliferation and thus act as growth factors. We have shown that both T3 and T4 exert potent pro-angiogenic effects in the human dermal microvascular endothelial cell assays and in the chick chorioallantoic membrane (CAM) and Matrigel model of angiogenesis. Taken together, these findings suggest that antagonist for thyroid hormones and/or related signaling pathways may represent compelling targets to treat cancer. Recent studies from our laboratory have shown that tetraiodothyroacetic acid (tetrac), a deaminated thyroid hormone analog, capable of preventing the binding of T4 to 1vB3 at the cell membrane exerted profound inhibitory effects on cellular proliferation and angiogenesis. The finding that tetrac inhibits cancer cell proliferation prompted us to investigate whether it may also stimulate cellular response to stress and thus reverse the development of drug resistance. Our results demonstrate that tetrac impacts on drug resistance and implicate at least two pathways that play key roles in cellular response to chemotherapy. In this Exploratory Application we will investigate the potential use of tetrac as a therapeutic tool for the treatment of cancer, particularly with regards to its possible role in suppressing the development of resistance to chemotherapy. The goal is to determine whether the anti-angiogenic and anti-proliferative actions of tetrac are initiated at the plasma membrane and not at the genomic level. To this end, we have generated PLGA nanoparticles covalently - linked to Tetrac on their surfaces. Preliminary data have shown that upon incubation with cancer cells, particles are preferentially localized to the plasma membrane. Hypothesis: Preferential targeting of Tetrac to the plasma membrane using Tetrac covalently-linked to nanoparticles (TclNPs) will provide a unique approach to study plasma membrane receptor-mediated actions of this hormone antagonist. In the context of the proposed study, we will investigate its anti- angiogenic and anti-proliferative actions. Because thyroid agents used systemically could potentially impact on multiple physiological pathways, the selective targeting of Tetrac to receptor(s) at the plasma membrane may be sufficient to significantly inhibit proliferation of aggressive tumors with fewer side effects. The use of biodegradable TclNPs loaded with a chemotherapeutic agent Paclitaxel can also be used for a controlled release of this drug at the tumor site and thus, is expected to result in better anti-tumor activity with reduced Paclitaxel toxicity. Specific Aims: 1. Experiments will be performed to evaluate the functional activities of TclNPs in vitro in MCF7 cells and in endothelial cells. Preliminary data using fluorescence microscopy showed that fluorescein-loaded TclNPs localize preferentially at the plasma membrane of the breast cancer MCF7 cells. Further studies will be performed to determine binding characteristics of TclNPs, and their effects on proliferation and cell migration. The effects of TclNPs also will be tested on pathways associated with the development or reversal of drug resistance in tumor cells. The effect of TclNPs will be compared to those of free Tetrac in these systems. 2. Pilot experiments to determine the optimum formulations of TclNPs will be performed in ova in the chick chorioallantoic membrane (CAM) tumor implant model of tumor growth and angiogenesis prior to performance of nanoparticle-targeted treatments of nude mice. The CAM model permits in ova pre-screening for bioactivity while limiting the use of more sentient and costly murine species. Nanoparticle formulations that show optimum anti-tumor and anti-angiogenesis activity in the CAM model will be tested in the orthotopic breast cancer model described in Experimental Methods. 3. Female athymic mice will have drug-resistant MCF7 human breast cancer cells implanted orthotopically into the fourth mammary gland. We will evaluate the effectiveness of targeted nanoparticles TclNPs, in reducing tumor growth and tumor angiogenesis. We will evaluate the efficacy of these formulations in limiting potential toxicities associated with systemic Tetrac administration and determine whether TclNPs, with or without Paclitaxel act to limit the development of drug resistance, as suggested by our preliminary in vitro studies. The novel nanoparticle system to be evaluated in these studies combines the properties of targeting and anti-tumor activities, and because Tetrac, Paclitaxel and PLGA nanoparticle systems are all approved for use in patients, could find potential clinical application in the foreseeable future. PUBLIC HEALTH RELEVANCE: Cancer cells have the unique ability to develop resistance to chemotherapeutic drugs, and so research on ways to reverse this phenomenon would have significant value in the treatment of cancer patients. This project will use a combination of two drugs, one of which impairs the cancer cell's ability to develop drug resistance, in a mouse model of breast cancer. A novel technology, the use of nanoparticles to encapsulate the test drugs, and direct them to tumors will be tested to determine whether these nanoparticles can improve the delivery of drugs and minimize the associated toxicities.